Adrenal chromaffin cells contain multiple hormones: epinephrine, norepinephrine, and enkephalins, each capable of producing unique physiological effects. Recently we have begun to investigate mechanisms which determine hormonal output from chromaffin cells. We have shown that the enhanced hormone release involves changes in mRNA levels of proenkephalin (pEK) and catecholamine synthetizing enzymes: tyrosine hydroxylase (TH) and phenylethanolamine-N- methyltransferase (PNMT). The purposes of this project were: 1) to examine nature of extracellular and intracellular signals involved in the controls of TH, PNMT and pEK mRNA levels; 2) to examine role of transcription and post-transcriptional mechanisms in these processes; 3) to determine whether expression of TH, PNMT and pEK genes could be differentially regulated. To address these questions we have used 3 experimental models: 1) in situ electrical stimulation of decentralized splanchnic nerves; 2) hypophysectomy and systemic administration of glucocorticoids; 3) primary culture of bovine adrenomedullary chromaffin cells. TH, PNMT and pEK mRNA levels were found differentially regulated by hormonal and neuronal inputs, by adenylate cyclase and intracellular catecholamines. We are currently investigating whether changes in mRNA levels are produced by transcriptional or posttranscriptional mechanisms, and whether differential regulations of individual mRNAs may provide bases for selective control of hormonal output from chromaffin cells. In the future, we intend to explore the role of different transmitters, hormones, growth factors, and second messenger systems in the control of TH, PNMT and pEK mRNA levels. To study coregulation of the expression of TH, PNMT, and pEK genes in individual cells we are currently developing in situ hybridization assay. We also plan to obtain clones of PNMT gene to further examine mechanisms which control transcription and stability of its products.